We have found that the RNA polymerase II transcription termination regions of genes from such diverse species as chickens, rabbits, and mice possess a highly conserved 20-nucleotide consensus sequence that we hypothesize is an important genetic element defining the 3' boundaries of vertebrate transcription units. To test this hypothesis we propose to determine if this sequence element is commonly observed at the 3' ends of a large number of eukaryotic structural genes. This will be done by using oligonucleotides which correspond to the consensus sequence element as hybridization probes to identify potential transcription termination sites associated with a number of other vertebrate structural gene clones. Transcription unit mapping will be used to determine if transcription terminates in restriction fragments which contain a copy of the 20-nucleotide consensus sequence element. Such a correlation would provide strong circumstantial evidence for a role of the consensus element in the process of transcription termination. In vivo functional tests of transcription termination, based on the use of recombinant adenovirus vectors or SV40-based replication vectors, will be developed to determine if the 20-nucleotide sequence element plays a functional role in the process of transcription termination. These studies will focus particularly on a functional analysis of the mouse beta-major globin and the dihydrofolate reductase terminator regions. Our analysis of the amplified dihydrofolate reductase domain has revealed evidence for the existence of three divergent transcription units located immediately in front of the dihydrofolate reductase gene: two in tandem and one embedded within another. This surprising model of transcription unit distribution and organizational arrangement will be rigorously tested for the dihydrofolate reductase locus. The generality of these features will be tested by an examination of transcription unit organization in the vicinity of the hamster CAD gene and the mouse adenosine deaminase gene. These studies should provide us with a better understanding of the distribution and organizational arrangement of transcription units in the mammalian genome.